2-Aminopurine modified DNA probe for rapid and sensitive detection of l-cysteine

Abstract A rapid, cost-effective and quencher-free fluorescence-based analytical method for sensitive detection of l -cysteine (Cys) based on 2-aminopurine (2-AP) labeled DNA probe and exonuclease I (Exo I) activity was developed. 2-AP labeled DNA probe includes two thymine (T)-T mismatches, which can bind with Hg2+ to form T-Hg2+-T pairing bases, resulting in stable hairpin with five base pairs in its stem. The target Cys can remove Hg2+ from the stem of the hairpin probe based on the high affinity of Cys with Hg2+, leading to the unfolding of the hairpin probe. At last, by adding Exo I, the resulted single-stranded DNA (ssDNA) will be digested to release free 2-AP with strong fluorescence. Under the optimal conditions, the sensing system exhibited a good and wider linear range from 0.4 to 400 nM (R2 = 0.997) and a detection limit as low as 0.16 nM for Cys. Furthermore, other amino acids without reductive sulfur group did not generate obvious change in fluorescence signals. Finally, the sensor can be used in diluted real samples with a good recovery rate, showing promising application in food, environmental and medical analysis.